TRANSCRIPTION FACTORS REGULATING CHORION GENE PROMOTERS

调节绒毛膜基因启动子的转录因子

• 批准号: 3468136
• 负责人: S ALEX MITSIALIS
• 金额: $ 12.63万
• 依托单位: BOSTON UNIVERSITY MEDICAL CENTER HOSP
• 依托单位国家: 美国
• 财政年份: 1990
• 资助国家: 美国
• 起止时间: 1990-07-01 至 1995-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3468136
• 关键词: Bombycidae; DNA binding protein; DNA footprinting; Drosophilidae; biochemical evolution; chorioallantoic membrane; computer assisted sequence analysis; developmental genetics; gene expression; genetic library; genetic promoter element; genetic regulatory element; genetically modified animals; histogenesis; nonmammalian vertebrate embryology; site directed mutagenesis; tissue /cell culture; transcription factor; transfection

The goal of the proposed research is the elucidation of mechanisms determining the tissue and temporal specificity of transcriptional activation. The model system to be used is insect choriogenesis, a process involving stringent development controls on the expression of a number of related genes, each of which is differentially regulated at eh transcriptional level. The studies will focus on the molecular biology of DNA-protein interactions regulating the expression of chorion genes and the analysis of the extent of conservation of such regulatory interactions between species. Specific aims will be: Localization and functional analysis of silkmoth chorion gene cis- regulatory elements. Regions on chorion gene promoters recognized in vitro by Drosophila follicular cell nuclear proteins and cloned Drosophila transcription factors will be identified through DNase I protection and methylation interference studies. These regions will be the targets of in vitro mutagenesis and the effect of these mutations on the tissue and temporal specificity of promoter activation will be analyzed in transgenic Drosophila. Cloning and characterization of silkmoth genes regulating chorion gene expression. Silkmoth follicular cell cDNA expression libraries will be screened for binding to synthetic oligonucleotides representing the recognition sites of follicular nuclear proteins. In addition, cDNAs of Drosophila transcription factors associated with chorion gene transcription will be used in sequence homology screening. Clones representing cDNAs of putative regulatory genes will be structurally characterized and used for the production of proteins in bacterial or tissue culture expression systems. Reconstruction of transcriptional control interactions in cell-free systems. Transcriptionally active nuclear extracts derived from follicles and tissue culture cell lines will be supplemented with Drosophila and silkmoth putative regulatory proteins and the effect on the in vitro transcription of chorion and non-chorion gene promoters will be analyzed. The interspecies supplementation studies will be especially revealing on the extent of functional conservation in transcriptional regulatory interactions. The above proposed studies will provide information on molecular mechanisms involved in transcriptional regulation of a class of genes under stringent developmental control. Interspecies conservation of regulatory interactions, such as observed in this insect system, are also evident in mammalian systems, although extensively characterized in only a few cases. Basic information on the extent of conservation of transcriptional regulatory interactions between species could be relevant in the development of animal models for human disease.

拟议研究的目标是阐明机制 确定转录的组织和时间特异性 激活。 要使用的模型系统是昆虫绒毛膜发生，这是一个过程 涉及对许多表达的严格开发控制 相关基因，每个基因在 eh 处受到差异调节 转录水平。 研究将集中于分子生物学 DNA-蛋白质相互作用调节绒毛膜基因的表达和 分析此类监管相互作用的保护程度 物种之间。 具体目标是： 蚕绒毛膜基因顺式定位及功能分析 监管要素。 体外识别的绒毛膜基因启动子区域 果蝇滤泡细胞核蛋白和克隆果蝇 转录因子将通过 DNase I 保护来识别 甲基化干扰研究。 这些地区将成为 体外诱变以及这些突变对组织和细胞的影响 将在转基因中分析启动子激活的时间特异性 果蝇。 蚕绒毛膜基因调控基因的克隆与表征 表达。 蚕滤泡细胞 cDNA 表达文库 筛选与代表的合成寡核苷酸的结合 滤泡核蛋白的识别位点。 此外，cDNA 果蝇转录因子与绒毛膜基因转录相关 将用于序列同源性筛选。 代表 cDNA 的克隆 假定的调控基因将被结构表征并用于 在细菌或组织培养表达中产生蛋白质 系统。 无细胞转录控制相互作用的重建 系统。 来自卵泡的转录活性核提取物 组织培养细胞系将补充果蝇和 蚕蛾假定的调节蛋白及其对体外的影响 将分析绒毛膜和非绒毛膜基因启动子的转录。 种间补充研究将特别揭示 转录调控中功能保守的程度 互动。 上述提议的研究将提供分子机制的信息 参与严格条件下一类基因的转录调控 发育控制。 种间保护监管 相互作用，例如在该昆虫系统中观察到的，在以下情况中也很明显： 哺乳动物系统，尽管仅在少数情况下得到广泛表征。 转录保守程度的基本信息 物种之间的监管相互作用可能与 开发人类疾病的动物模型。